Herein, we present a poly-adenine (polyA)-mediated programmably engineered FRET-nanoflare for ratiometric intracellular ATP imaging with anti-interference capability. The programmable polyA attachment is advantageous in enhancing the signal response for ATP. Moreover, the FRET-based nanoflare is capable of avoiding false-positive signals due to probe degradation in a complex environment, which has great potential for clinical diagnosis.